1. Field of the Invention
The present invention relates to the field of tyrosinase inhibitors and to methods and compositions of treatment involving inhibition of this enzyme.
2. Related Art
Presented below is background information on certain aspects of the present invention as they may relate to technical features referred to in the detailed description, but not necessarily described in detail. That is, individual parts or methods used in the present invention may be described in greater detail in the materials discussed below, which materials may provide further guidance to those skilled in the art for making or using certain aspects of the present invention as claimed. The discussion below should not be construed as an admission as to the relevance of the information to any claims herein or the prior art effect of the material described.
The present invention relates to novel biological agents, specifically oligopeptides that reduce the enzymatic activity of tyrosinase. These agents have use as research and development tools in basic science investigation, in diagnostic applications, as cosmeceuticals for the treatment of pigmentary disorders including skin conditions characterized by hyperpigmentation, and as therapeutics for the treatment of pathological conditions that rely on tyrosinase enzyme activity to promote their tumorigenicity or other adverse effects.
Melanin plays an important role in protecting the human body from the harmful effects of ultraviolet rays. Melanin is also an important factor in medical science and cosmetology. It is known that melanin is formed or synthesized in skin tissues. Excessive amounts of melanin darken the skin, and the nonuniform distribution of melanin causes chloasma and ephelis, both of which are skin disorders.
Melanin is a photoprotective biopolymer synthesized by melanocytes in discrete organelles known as melanosomes. Once filled with melanin, epidermal melanosomes are transferred to keratinocytes, where they form a supranuclear cap to protect DNA against ultraviolet irradiation. The type, amount, and distribution pattern within surrounding keratinocytes help determine skin color. Melanin forms through a series of oxidative reactions involving the amino acid tyrosine and the enzyme tyrosinase. Since its discovery in 1896 by Bourquelot and Bertrand, the enzyme tyrosinase has been extensively studied and its rate limiting role in the melanin biosynthetic pathway has since been elucidated through the work of Raper et al (Biochem J., 1923, 17: 454-469), Mason H S (J. Biol. Chem., 1948, 172: 83-99), Cooksey et al (J. Biol. Chem., 1997, 272: 26226-26235) and Schallreuter et al (Exp. Dermatol., 2008, 17: 395-404).
Tyrosinase is a melanocyte-specific, multi-functional, glycosylated, copper containing oxidase with a molecular weight of approximately 60-70 kDa in mammals. Defects in the enzyme's activity lead to albinism in humans. It is believed that racial differences in skin color may primarily be due to differences in tyrosinase activity. Melanocytes derived from African skin demonstrate up to ten times more activity and melanin production than melanocytes from Caucasian skin. However, this is not due to a greater abundance of tyrosinase, as both skin types have been shown to contain a similar density of tyrosinase molecules. The first two steps in the melanogenic pathway are hydroxylation of L-tyrosine to 3-4-dihydroxyphenylalanine (L-dopa) and the oxidation of L-dopa to o-dopaquinone, a highly reactive compound that spontaneously polymerizes to form melanin.
The most common pigmentary disorders for which patients seek treatment are melasma and post-inflammatory hyperpigmentation (Lynde, C. B., Kraft, J. N., and Lynde, C. W. (2006) Skin Therapy Lett 11, 1-6). These conditions may have a major impact on a person's psychological and social well-being, contributing to lower productivity, social functioning, and self-esteem (Finlay, A. Y. (1997) Br J Dermatol 136, 305-314) as well as a negative impact on the person's health-related quality of life (Balkrishnan, R., McMichael, A. J., Camacho, F. T., Saltzberg, F., Housman, T. S., Grummer, S., Feldman, S. R., and Chren, M. M. (2003) Br J Dermatol 149, 572-577; Taylor, A., Pawaskar, M., Taylor, S. L., Balkrishnan, R., and Feldman, S. R. (2008) J Cosmet Dermatol 7, 164-168). Thus, pharmacological agents that can reduce hyperpigmentation are of great interest clinically (Solano, F., Briganti, S., Picardo, M., and Ghanem, G. (2006) Pigment Cell Res 19, 550-571), and include hydroquinone (dihydroxy-benzene; HQ) (Jimbow, K., Obata, H., Pathak, M. A., and Fitzpatrick, T. B. (1974) J Invest Dermatol 62, 436-449), arbutin (Maeda, K., and Fukuda, M. (1996) J Pharmacol Exp Ther 276, 765-769), kojic acid (Cabanes, J., Chazarra, S., and Garcia-Carmona, F. (1994) J Pharm Pharmacol 46, 982-985), vitamin C (Kojima, S., Yamaguchi, H., Morita, K., and Ueno, Y. (1995) Biol Pharm Bull 18, 1076-1080), retinol (Pathak, M. A., Fitzpatrick, T. B., and Kraus, E. W. (1986) J Am Acad Dermatol 15, 894-899), azelaic acid (Schallreuter, K. U., and Wood, J. W. (1990) Arch Dermatol Res 282, 168-171), as well as a number of botanical compounds (Hwang, J. H., and Lee, B. M. (2007) J Toxicol Environ Health A 70, 393-407).
To date, the mainstay treatment for hyper-pigmentation has been hydroquinone (HQ) at a concentration of 2-4% (Ennes, S., et al. Journal of Dermatological Treatment, 2000, vol 11, pages 173-179). In addition to tyrosinase inhibition, HQ has been shown to degrade melanosomes and induce apoptosis in melanocytes (Arck, P. C., et al. FASEB Journal, 2006, vol 20, pages 1567-1569; Inayat-Husain, S. H., et al. Chem. Res. Toxicol., 2005, vol 18, pages 420-427; and Penney, K. B., et al. J. Invest. Dermatol, 1984, vol 82, pages 308-310). It was also found that HQ achieves its hypopigmenting effects through depletion of glutathione, leading to the generation of reactive oxygen species and subsequent oxidative damage of membrane lipids and proteins (Briganti, S., et al., Pigment Cell Research, 2003, vol 16, pages 101-110). HQ's pleiotropic effects are complicated by a number of adverse reactions including contact dermatitis, irritation, transient erythema, burning, prickling sensation, leukoderma, chestnut spots on the nails, hypochromia and ochronosis (Engasser, P. G., J. Am. Acad. Dermatol., 1984, vol 10, pages 1072-1073; Romaguera, C., et al. Contact Dermatitis, 1985, vol 12, page 183). Moreover, HQ is potentially mutagenic to mammalian cells. HQ and its derivative arbutin are both catabolized to benzene metabolites with the potential for bone marrow toxicity (Curto, E. V., et al. Biochem. Pharmacol., 1999, vol 57, pages 663-672; Zhou, H., et al. Mol. Pharmacol., 2009, vol 76, pages 579-587).
Hydroquinone (HQ) has been used since the 1950's in commercially available over-the-counter skin lightener products and since the 1960's as a commercially available medical product. It is also used in cosmetic products such as hair dyes and products for coating fingernails. However, beginning in 2001, HQ is no longer authorized for use in cosmetic skin lightening formulations in European Union countries, although products containing arbutin, an analogue of HQ, and botanicals, including plants that naturally contain HQ and arbutin, continue to remain available in European countries See also, Matsubayashi et al., “Pharmaceutical and clinical assessment of hydroquinone ointment prepared by extemporaneous nonsterile compounding,” Biol Pharm Bull. 2002 January; 25(1):92-6. As disclosed there, ointments of the skin depigmentation agent HQ have been prepared by extemporaneous nonsterile compounding in Japan by imitating skin lightening creams commercially available in the U.S.A. and European Union. However, various problems have been observed including chromatic aberration of HQ ointments, relatively large variability of efficacy, and undesirable side effects although they were mild. HQ has a published IC50 of about 700 μM.
Therapies containing HQ have been outlawed in Asian countries, making the standard HQ treatment inaccessible to a large number of people suffering from this condition. In fact, the United States FDA has issued a notice indicating that it may too ban the use of HQ domestically. Furthermore, HQ has been associated with visceral malignancy and long-term topical delivery may be a potentially harmful therapeutic option. HQ in the best of circumstances leads to only a partial alleviation of hyperpigmentation. Some cosmeceutical formulations (i.e. compositions) have included other active ingredients such as kojic acid, arbutin, and vitamin C but efficacy has thus far been disappointing due to problems with chemical instability or inability to deliver the active to the appropriate layer of skin. HQ and its derivative arbutin are both catabolized to benzene metabolites with the potential for bone marrow toxicity (Zhou, H., Kepa, J. K., Siegel, D., Miura, S., Hiraki, Y., and Ross, D. (2009) Mol Pharmacol 76, 579-587). Similarly, kojic acid, fungally derived HQ derivative, has been shown to promote thyroid and liver carcinogenicity in rodent models (Fujimoto, N., Onodera, H., Mitsumori, K., Tamura, T., Maruyama, S., and Ito, A. (1999) Carcinogenesis 20, 1567-1571), leading to its ban in Switzerland. Although higher concentrations have been utilized, patients often discontinue treatment due to skin irritation. This led to the addition of topical steroids in order to reduce irritation from the active ingredients such as retin A and HQ. Since melasma and other hyperpigmentary disorders often take months to years to treat, use of topical steroids on the face at the strength required to combat irritant effects of active ingredients is not possible without causing topical steroid-induced side effects. When medium or greater potency topical steroids are used on the face for more than several weeks consecutively, skin atrophy, fragility and telengiectasia commonly occur. This side effect profile is unacceptable, especially in areas such as the face.
Infrared lasers have been used with some success. They generally are more effective for conditions that localize pigment to the deeper skin areas such as the dermis. In order to effectively treat the epidermis, an ablative treatment is usually employed. This therapy is associated with significant downtime for the patient, including creation of second-degree burn or erosion leaving the patient susceptible to infection. In addition, laser therapy is an expensive treatment option that many patients cannot afford. In extreme cases, depigmentation of the skin has been elected when bleaching agents have been unsuccessful. Numerous pathological conditions can lead to the deposition of pigment into the skin aberrantly. For example, it is well known that hormonal imbalance can cause facial and extremity hyperpigmentation, most frequently observed in women during or following pregnancy. Often times, this hyperpigmentation becomes aesthetically disfiguring, leading to problems with self-esteem and embarrassment in social situations. Melasma often times affects individuals with Fitzpatrick type IV-VI skin. This constitutes a significant portion of the worldwide population.
A large number of individuals with Fitzpatrick type IV to VI skin are of Asian descent.
According to the Fitzpatrick skin type scale, based on a test of appearance and skin reaction to sun exposure, individuals are generally categorized as follows:
Type I: Very fair skin tone, blond or redhead,
Type II: Light skin tone, will tan, but usually burns.
Type III: White to olive skin tone, sometimes burns.
Type IV: Medium brown skin tone, rarely burns.
Type V: Dark brown skin tone, very rarely burns.
Type VI: Black skin tone, very dark eyes, burn resistant.
In addition to melasma, hyperpigmentation of aesthetically sensitive locations such as the face may take place after inflammation due to disorders such as acne or rosacea, amongst others. These conditions may also lead to significant psychological discomfort. In the United States, $13 billion are spent on cosmeceuticals each year. With the anticipated ban of HQ in the US market, in conjunction with the continued stability (vitamin C) or delivery (arbutin, kojic acid, etc) problems of other non-pharmacological agents currently on the market, oligopeptide inhibitors may provide a solution to this large unmet need.
Specific Patents and Publications
Scot et al., “Production of cyclic peptides and proteins in vivo,” Proc. Nat. Acad. Sci. Vol. 96, Issue 24, 13638-13643, Nov. 23, 1999, discloses the production of the cyclic, eight-amino acid tyrosinase inhibitor pseudostellarin F in bacteria.
Verma et al., “Modulation of agonist binding to human dopamine receptor subtypes by L-prolyl-L-leucyl-glycinamide and a peptidomimetic analog,” J Pharmacol Exp Ther. 2005 December; 315(3):1228-36. Epub 2005 Aug. 26, discloses the role of the hypothalamic tripeptide L-prolyl-L-leucyl-glycinamide (PLG) and its conformationally constrained analog 3(R)-[(2(S)-pyrrolidinylcarbonyl) amino]-2-oxo-1-pyrrolidineacetamide (PAOPA) in modulating agonist binding to human dopamine (DA) receptor subtypes.
U.S. Pat. No. 6,165,982 to Yamada, et al., issued Dec. 26, 2000 entitled “Use of sericin as antioxidants and tyrosinase inhibitors,” discloses a composition useful as an antioxidant or an inhibitor for tyrosinase activity which comprises as an active ingredient a sufficient amount of sericin to exert an antioxidizing ability. Sericin is a high molecular weight, natural, soluble glycoprotein constituent of silk. Sericin binds to the keratin of skin and hair, forming a protective film.
U.S. Pat. No. 5,126,327 to Takeuchi, et al., issued Jun. 30, 1992, entitled “Melanocyte-stimulating hormone inhibitor and external preparation containing the same,” discloses a melanocyte-stimulating hormone inhibitor which has certain amino acid sequences, an acyl group having 1 to 12 carbon atoms, an amino acid residue, or acylated derivative thereof having 1 to 12 carbon atoms, peptide residue having 2 to 40 amino acid residues or acylated derivative thereof.
U.S. Pat. No. 7,025,957 to Arquette, issued Apr. 11, 2006, entitled “Composition and method to whiten skin,” discloses a composition effective as a skin whitening agent. The composition includes Simmondsin, which is a glycoside extracted from jojoba meal (Simmondsia chinensis). In certain embodiments, the composition comprises an extract of jojoba (Simmondsia chinensis). The composition is administered by topically applying to an individual a formulation in an amount effective to whiten skin, where that composition comprises a jojoba extract.
U.S. Pat. No. 7,083,781 to Fotinos, et al., issued Aug. 1, 2006, entitled “Film forming polymers, methods of use, and devices and applications thereof,” discloses compositions and methods for delivering active agents to the skin of a subject, including a polymer, an active ingredient and a solvent, the compositions being capable of delivery by rolling, spreading, aerosol or in droplets and of forming a film in contact with the skin. A cosmetic active agent known in the art may be incorporated in the film forming compositions for improving skin appearance. Anti-hyperpigmentation agents typically used for counterbalancing this condition can include tyrosinase inhibitors such as peptide mixtures and plant extracts, fermentation products, and antioxidants such as hydroquinone, kojic acid, ascorbic acid derivatives, synthetic or natural derivatives of hydroquinone and hydroquinone precursors. In preferred embodiments of the invention, anti-hyper pigmentation agents are Melawhite of Pentharm Ltd., Basel, Switzerland; Biowhite™ of Coletica, France; Etioline of Sederma, France; Arbossa of Kelesima, Italy; Gatuline whitening of Gattefosse, France; Ascorbocilan C of Exsymol, Monaco; and Kojic acid of Alps Pharm., Japan.
U.S. Pat. No. 7,125,572 to Lee, issued Oct. 24, 2006, entitled “Tyrosinase inhibitor extract,” discloses a tyrosinase inhibitor extract from lemon peels. The tyrosinase inhibitor provides advantageous skin whitening effects. According to the invention, the tyrosinase inhibitor extract of the invention has a main absorbance at 280 nm. This indicates that the tyrosinase inhibitor extract contains a protein or peptide. It is believed that the protein or peptide is the main active component for inhibiting tyrosinase. The other components of the extract may provide additional effects such as anti-aging and anti-oxidation. The tyrosinase inhibitor extract can be prepared to be in various forms, including lotions, emulsions, creams, ointments, sticks, solutions, packs, and gel. The tyrosinase inhibitor extract may be admixed with any ingredients ordinarily used in cosmetics, such as oily substances, humectants, thickeners, preservatives, emulsifiers, medical ingredients, perfumes, emulsification stabilizers and the like.